P1253: REDUCED EXPRESSION OF THE CHEMOKINES CXCL2, CXCL3 AND CXCL8 PROVIDES A POTENTIAL MECHANISM PROMOTING IMMUNE EVASION FACILITATING AML RELAPSE FOLLOWING ALLOGENEIC HSCT
Authors: Sarah Charrot, Jennifer Ball, Findlay Copley, James Aries, Matthew Mee, Symeon Theocharidis, James Heward, John Gribben, Jeff Davies
Journal: HemaSphere (2023)

Topic: 21. Stem cell transplantation - Experimental Background: 20-50% of patients treated with allogeneic hematopoietic stem cell transplantation (AHSCT) for acute myeloid leukemia (AML) subsequently relapse. Relapse is associated with immune evasion from the alloreactive graft-versus-leukaemia (GvL) effect by either downregulation of MHC class II or upregulation of inhibitory molecules. The chemokines CXCL2, CXCL3 and CXCL8 mediate their actions through the CXCR2 receptor and are known to have a role in tumour immunogenicity, but little is known about their role at AML relapse post-AHSCT. Aims: To interrogate gene expression profiles of matched AML samples taken at diagnosis and post-AHSCT relapse in order to identify cell intrinsic mechanisms promoting immune escape, and assess the functional impact of the dysregulated pathways on allogeneic immune responses. Methods: Nine patients with relapsed AML post-AHSCT were included in the discovery cohort, of which six had paired diagnosis and relapse samples. All patients were treated on the same reduced intensity, T-replete platform and all achieved full donor T-cell chimerism. AML blasts were isolated by fluorescence activated cell sorting prior to nucleic acid extraction and assessment of immunogenicity. Gene expression profiling was carried out by RNAseq and evaluation of accessible chromatin by ATACseq. Susceptibility to T cell-mediated killing was assessed in allogeneic co-culture with mismatched healthy donor peripheral blood mononuclear cells (PBMCs). The functional impact of the CXCR2 pathway was investigated in both primary AML samples and a panel of AML cell lines in a co-culture model of alloreactivity (Fig.1a). Results: Paired samples from four patients were assessed for their capacity to stimulate an in vitro allogeneic T cell response at relapse compared to diagnosis, of which three showed reduced ability to stimulate both CD4 and CD8 T cell proliferation, suggesting that immune evasion was a factor in relapse in this cohort (Fig.1b). Gene expression analysis demonstrated a high degree of correlation between diagnosis and relapse samples (Fig.1c), with only 22 significantly differentially expressed genes, including downregulation of the chemokines CXCL2, CXCL3 and CXCL8 (Fig.1d,e). Interrogation of paired diagnosis and post-chemotherapy relapse data in the OHSU AML cohort showed that none of these genes were significantly differentially expressed at relapse following chemotherapy, suggesting that the findings were post-AHSCT relapse specific. The importance of CXCL8 was further emphasised by pathway analysis of differentially accessible chromatin, which showed significant enrichment of regulation of CXCL8 secretion pathways. Finally, the roles of these chemokines were investigated in an in vitro model of alloreactivity. Baseline gene expression of the differentially expressed chemokines stratified a panel of AML cell lines into those which were susceptible to allogeneic T cell-mediated killing in co-culture and those which were resistant (Fig.1f). Co-culture was performed between AML blast stimulators from post-AHSCT relapse samples and healthy donor PBMC responders. Both CXCL2 and CXCL8 treatment led to an increase in CD4 T cell proliferation and in PD-1 expression (Fig.1g). Conversely, CXCR2 inhibition with AZD5069 led to reduced CD4 proliferation and PD-1 expression. Summary/Conclusion: This study further highlights the importance of immune evasion in mediating AML relapse following AHSCT. The chemokines CXCL2 and CXCL8 were identified as potential mediators of this process and may provide prospective targets to enhance the GvL effect to prevent or treat post-AHSCT relapse.Keywords: Chemokine, Acute myeloid leukemia, Allogeneic hematopoietic stem cell transplant